Time-Released Black Phosphorus Hydrogel Accelerates Myocardial Repairing through Antioxidant and Motivates Macrophage Polarization Properties

Figures

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  Fig. 1.. Schematic diagram of synthesis and action mechanism of CS–Gel–BP@PDA.

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  Fig. 2.. Characterization of BPNSs and BP@PDA. (A) Transmission electron microscopy of BPNSs. (B) Transmission electron microscopy of BP@PDA. (C) Atomic force microscopy of BPNSs and (D) thickness measurements. (E) Detection of XRD diffraction peaks of BPNSs and BP@PDA, compared with the standard black phosphorus block. (F) Absorption peaks of Raman spectra of BPNSs and BP@PDA. (G) Zeta potential detection of BPNSs and BP@PDA. (H) FTIR of BPNSs, PDA, and BP@PDA. (I) UV-Vis detection of BPNSs, PDA, and BP@PDA. (J) STEM image and STEM-EDX elemental mapping images of BP@PDA.

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  Fig. 3.. Characterization of the composite hydrogel. Representative scanning electron microscopy images of (A) CS-Gel and (B) CS-Gel-BP@PDA. (C) Rheological property test of CS-Gel and CS-Gel-BP@PDA. (D) Tensile-strain curves of CS-Gel and CS-Gel-BP@PDA. (E) Compression-strain curves of CS-Gel and CS-Gel-BP@PDA. (F) Dissolution performance test of CS-Gel and CS-Gel-BP@PDA. (G) Digital photographs of natural degradability of CS-Gel-BP@PDA. (H) Digital photographs of the degradability of BP@PDA. (I) UV-Vis spectra of supernatants of BP@PDA in H2O2 solution on days 0, 3, 5, 7, and 14. (J) X-ray photoelectron spectroscopy detection of supernatants of BP@PDA in H2O2 solution on day 0 and (K) day 14. (L) Composite hydrogel P-element release capacity curves. (M) DPPH radical scavenging ability of CS-Gel with different concentrations of CS-Gel-BP@PDA. (N) ABTS radical scavenging ability of CS-Gel with different concentrations of CS-Gel-BP@PDA. (Scale bar: 10 µm. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001)

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  Fig. 4.. Antioxidant capacity of the composite hydrogel in vitro. (A) CCK-8 was used to detect the relative activity of H9C2 in different treatment groups. (B) Calcein-AM staining on CS–Gel and CS–Gel–BP@PDA after 24 h of culture. (C) DHE fluorescence staining and quantitative statistical analysis of different treatment groups. (D) DCFH-DA fluorescence staining and quantitative statistical analysis of different treatment groups. (E) JC-1 fluorescent staining of different treatment groups. (Scale bar: 50 µm. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001)

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  Fig. 5.. The anti-inflammatory ability of composite hydrogel in vitro. (A) Immunofluorescence colocalization and quantitative statistical analysis of IL-10 and IL-6 in different treatment groups. (B) ELISA results of IL-6, IL-10, TNF-α, and TGF-β in the supernatant of RAW246.7 cell culture medium in different treatment groups. (Scale bar: 10 µm. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001) DAPI, 4′,6-diamidino-2-phenylindole.

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  Fig. 6.. CS–Gel–BP@PDA improved cardiac function and myocardial fibrosis in MI rats. (A) Timeline of experiments in rats treated with CS–Gel–BP@PDA composite hydrogel. (B) Electrocardiograms of rats before and after MI. (C) Echocardiography and (D) H&E, Masson staining, and quantitative statistics of cardiac sections in different treatment groups on day 28 after MI. (E) Quantitative analysis of EF, FS, LVIDD, and LVIDS in different treatment groups. (F) WB detection of α-SMA and Col I protein expression at infarct sites in different treatment groups. (G) Col III immunofluorescence staining and quantitative statistical analysis of infarct sites in different treatment groups. (*P < 0.05, **P < 0.01, ***P < 0.001, ****P< 0.0001) GAPDH, glyceraldehyde-3-phosphate dehydrogenase.

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  Fig. 7.. CS–Gel–BP@PDA removal of ROS in the infarcted microenvironment of rats with MI alleviated oxidative stress and promoted macrophage polarization to M2 phenotype. (A) Immunofluorescence staining of DHE, Caspase-3, TNF-α, and IL-10 at the infarct site after 28 d of modeling in each group. (B) Quantitative statistical analysis of DHE, caspase-3, TNF-α, and IL-10 immunofluorescence staining at the infarct site on day 28 of modeling in each group. (C) WB detection of NF-κB apoptosis pathway-related protein expression and (F) quantitative statistical analysis on day 28 of modeling in each group. (D) Immunofluorescence co-localization staining of CD206/CD86 in the infarcts on day 28 of modeling in each group. (E) WB detection of macrophage polarization-related protein expression and (G) quantitative statistical analysis on day 28 of modeling in each group. (Scale bar: 50 µm. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001)

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  Fig. 8.. System toxicity. (A) Systemic toxicity evaluation. H&E staining of liver, lung, spleen, and kidney in Sham, Saline, CS-Gel, and CS-Gel-BP@PDA groups on day 28 after modeling. (B) Liver and kidney function (ALT, alanine aminotransferase; AST, aspartate aminotransferase; CREA, creatinine; UREA, urea) was evaluated in Sham, Saline, CS-Gel, and CS-Gel-BP@PDA groups on day 28 after modeling.